Amyloid beta-peptide (Abeta) is central to the pathogenesis of Alzheimer's disease (AD), and the AD brain is under extensive oxidative stress. Previous research from our laboratory combined these two observations into a model for neurodegeneration in AD, a model based on Abeta-associated free radical oxidative stress. In neurons, A-beta associated free radical oxidative stress and toxicity depend critically on methionine residue 35 of Abeta (1-42), and both are prevented or modulated by exogenous antioxidants. We now propose to extend these observations to gain insight into the mechanisms and role of Met-35 of Abeta (1-42) in the oxidative stress and neurotoxic properties of this peptide. In addition, we propose studies to gain insight into the mechanisms and sequelae of Abeta-associated free radical oxidative stress in AD brain and transgenic models of this disorder. In Specific Aim #1, we will test the hypothesis that the specific location of Met-35 is important in these properties of Abeta (1-42), and that Met is the initiating locus of Abeta-associated free radicals. Further, we will investigate whether interchain H-atom transfer occurs, as it has been proposed, between Gly-33 and a free radical on Met-35. Lastly, we will test the hypothesis that Met-35 is the initiating site of neuronal membrane lipid peroxidation induced by Abeta (1-42). In Specific Aim #2, we will test the hypothesis that transgenic models of familial AD, PS-1 mutant knock-ibn mice and doubly transgenic PS-1/APP mutant mice have increased markers of an increased vulnerability to oxidative stress. In Specific Aim #3, we will test the hypothesis that pharmacologically or genetically increased levels of antioxidants will modulate intrinsic or Abeta-induced oxidative stress in these two transgenic models of AD. In Specific Aim #4, we will employ novel methods to identify specifically oxidized proteins in involved AD brain regions compared to non-involved cerebellum as a function of early-, mid-, or late-stage AD. We expect to learn the importance of protein oxidation as an early event in AD. This is a comprehensive set of proposed studies, employing novel approaches in systems ranging from AD brain to transgenic animal models of AD, to neuronal cultures, to solution chemistry of Abeta, designed to gain insight into Abeta- associated free radical oxidative stress and neurotoxicity and their modulation by endogenous and exogenous antioxidants. The increased understanding that will result from these studies will provide insight into potential therapeutic interventions in this important dementing disorder that affects millions of Americans.